Glucagon is a 29-residue peptide hormone that regulates glycogenesis. The structure of glucagon may be represented as follows: ##STR1##
The abbreviations utilized herein are those recommended by IUPAC-IUB [See Eur. J. Biochemo 138, 9 (1984)].
The nomenclature used to define the peptides is that specified by Schroder & Lubke, "The Peptides", Academic Press (1965), wherein in accordance with conventional representation the amino group at the N-terminal appears to the left and the carboxyl group at the C-terminal to the right. Where the amino acid residue has isomeric forms, it is the L- or naturally occurring form of the amino acid that is represented unless otherwise expressly indicated.
Insulin, as is known, rapidly decreases elevated blood sugar. It is believed that, in humans, diabetes is only observed when insulin levels are low and glucagon levels are simultaneously elevated. The absence of insulin causes a rapid increase in blood glucose. Large amounts of insulin are required to reduce the glucose levels to normal. The maintenance of stable levels is difficult and subject to considerable fluctuation. This wide fluctuation is responsible, at least in part, for the clinical difficulties experienced in diabetes.
Glucagon appears to act by binding to liver membrane receptors thereby activating adenylate cyclase which, in turn, triggers a series of reactions including the production of cyclic adenosine monophosphate (cAMP), which activates phosphorylase and inhibits glycogen synthetase, thereby contributing to elevated glucose levels in the blood.
Recently, considerable effort has been expended to develop glucagon antagonists that will bind to liver membrane receptors but do not have the ability to transduce the signal to activate adenylate cyclase. One such product is N.alpha.-trinitrophenyl [12-homoarginine] glucagon. This product does bind to the glucagon receptor without significant activation of adenylate cyclase. It also activates another signaling system in the hepatocyte membrane leading to the production of inositol trisphosphate and calcium ions. A useful antagonist will block the action of endogenous glucagon by preventing it from binding to the liver membrane receptors and thereby producing cAMP and glucose in the cell, and the ultimate elevation of blood sugar. Such products would be useful to reduce a diabetic's need for injections or infusion of insulin.
An ideal glucagon antagonist would (1) be completely inactive toward stimulation of adenylate cyclase and production of cAMP, (2) bind as well as, or better than, glucagon itself to the liver membrane receptor, (3) compete with glucagon for receptor binding, (4) at moderate concentrations fully inhibit the action of glucagon toward the activation of adenylate cyclase, and (5) have a satisfactory inhibition index.
The inhibition index is the molar ratio of antagonist to agonist which reduces the biological response to 50% of the value in the absence of antagonist. It will be discussed more fully hereinafter.
U.S. Pat. Nos. 4,879,273 and 5,143,902 describe certain useful glucagon analogs in which the aspartic acid residue at the 9-position of the glucagon molecule is removed or replaced with another amino acid residue either in the D-form or the L-form. The replacement may be selected from any of a number of amino acids both natural and synthetic, including hydrophobic and hydrophilic amino acids, aliphatic amino acids, aryl amino acids, basic amino acids and acidic amino acids.
It was observed that compounds of the class described in these patents either with or without the histidine residue at the 1-position are useful adjuncts to insulin therapy in the control of blood glucose levels. The preferred compounds for such utility, as disclosed in the prior patents are:
des His.sup.1 [Gly.sup.9 ]glucagon SEQ ID NO: 2 PA1 des His.sup.1 [Nle.sup.9 ]glucagon SEQ ID NO: 3 PA1 des His.sup.1 [Lys.sup.9 ]glucagon SEQ ID NO: 4 PA1 des His.sup.1 [Glu.sup.9 ]glucagon SEQ ID NO: 5 PA1 des His.sup.1 [Glu.sup.9 Lys.sup.17,18 Glu.sup.21 ]glucagon SEQ ID NO: 6
and the corresponding carboxy terminal amides of such compounds.
It has now been discovered that the utility of the compounds of the previous patents for the treatment of diabetes can be remarkably and surprisingly improved by replacement of the serine residue at the 16-position and the optional replacement of the serine residue at the 11-position and/or the aspartic acid residue at the 21-position. However, the key to the utility of the compounds is still the 9-position.